Neuropharmacology 39 (2000) 1451–1462 www.elsevier.com/locate/neuropharm Cloning, expression and pharmacological characterisation of the mouse somatostatin sst 5 receptor Dominik Feuerbach, Dominique Fehlmann, Caroline Nunn 1 , Sandra Siehler 2 , Daniel Langenegger, Rochdi Bouhelal, Klaus Seuwen, Daniel Hoyer * Nervous System Research, S-386/745, Novartis Pharma AG, CH-4002 Basel, Switzerland Accepted 13 March 2000 Abstract The mouse somatostatin (somatotropin release inhibiting factor, SRIF) sst 5 receptor coding sequence was cloned from a mouse BALB/c genomic library. It shows 97% and 81% homology with the corresponding rat and human receptors, respectively. The msst 5 receptor messenger RNA (mRNA) is present at low levels in the adult mouse brain, with significant expression in a few nuclei only, e.g. in the septum (lateral septal nuclei) or the amygdala (medial amygdaloid nucleus); very few signals were observed in the mesencephalon, metencephalon, and myelencephalon (except the dorsal motor nucleus of the vagus nerve). The msst 5 receptor was stably expressed in the hamster fibroblast cell line CCL39-SRE-Luci, which harbours the luciferase reporter gene driven by the serum responsive element. [ 125 I]LTT-SRIF-28 ([Leu 8 , D-Trp 22 , 125 I-Tyr 25 ]-SRIF-28), [ 125 I]Tyr 10 -CST, [ 125 I]CGP 23996, and [ 125 I]Tyr 3 -octreotide labelled msst 5 receptors with high affinity (pK d values: 11.0, 10.15, 9.75 and 9.43) and in a saturable manner, but defined different Bmax values: 697, 495, 540 and 144 fmoles/mg, respectively. [ 125 I]LTT-SRIF-28- labelled sites displayed the following rank order: SRIF-28 rCST-14 somatuline  CGP-23996= SRIF-14= octreotide, whereas [ 125 I]Tyr 3 -octreotide-labelled sites displayed a different profile: octreotide  SRIF-28 rCST-14= somatuline  SRIF-14 CGP- 23996. The pharmacological profiles determined with [ 125 I]LTT-SRIF-28, [ 125 I]CGP 23996 and [ 125 I]Tyr 10 -CST correlated highly significantly (r 2 =0.88–0.99), whereas [ 125 I]Tyr 3 -octreotide binding was rather divergent (r 2 =0.77). Also, human and mouse sst 5 receptor profiles are very different, e.g. r 2 =0.385 for [ 125 I]Tyr 10 -CST and r 2 =0.323 for [ 125 I]LTT-SRIF-28-labelled sites. Somatostatin induces expression of luciferase reporter gene in CCL39-SRE-Luci cells. The profile was consistent with a msst 5 receptor-mediated effect although apparent potency in the luciferase assay was much reduced compared to radioligand binding data: Octreotide = SRIF-28 rCST-14= SRIF-14= CGP-23996. Octreotide, SRIF-28, BIM23052 and D Tyr Cyanamid 154806 behaved as full or nearly full agonists in comparison to SRIF-14, whereas the other compounds had relative efficacies of 40 to 70%. The present study shows that agonists radioligands define apparently different receptor populations in terms of number of sites and pharmacological profile in cells expressing a single recombinant receptor. These variations suggest that the conformation of the ligand receptor complex may vary depending on the agonist. Further, the msst 5 receptor, although primarily coupled to Gi/Go proteins, is able to stimulate luciferase gene expression driven by the serum responsive element. Finally, it is suggested that putative sst 2 selective agonists e.g. octreotide, RC160 or BIM23027 show similar or higher potency at msst 5 receptors than SRIF-14.  2000 Elsevier Science Ltd. All rights reserved. Keywords: Somatostatin (SRIF); Mouse somatostatin receptor 5 (msst 5 ); CCL39 Chinese hamster lung fibroblast cells; Serum responsive element (SRE); Luciferase; In situ hybridisation 1. Introduction The cyclic peptide somatostatin (SRIF, somatotropin release inhibiting factor) exists in two different forms, * Corresponding author. Tel.: + 41-61-324-4209; fax: + 41-61-324- 4866. E-mail address: daniel1.hoyer@pharma.novartis.com (D. Hoyer). 1 Present address: School of Biological Sciences, G 38 Stopford Building, Manchester University, Oxford Road, Manchester, M13 9PT, UK. 2 Present address: Department of Pharmacology, University of Pennsylvania, School of Medicine, 3620 Hamilton Walk (John Morgan Building), Philadelphia PA 19103, USA. 0028-3908/00/$ - see front matter  2000 Elsevier Science Ltd. All rights reserved. PII:S0028-3908(00)00063-0 namely with 14 amino acids (SRIF-14) and in an N- terminally extended form with 28 amino acids (SRIF- 28). Five different receptors for SRIF (sst 1 to sst 5 , Hoyer et al. 1994, 1995; Bell and Reisine, 1993) which all belong to the G-protein coupled receptor family have been cloned from various species including rat and man. They are grouped in two classes according to structure and pharmacology (Hoyer et al., 1995): sst 2 , sst 3 and sst 5 with high affinity for octreotide, seglitide and somatuline and sst1/sst4 with very low affinity for these short ana- logues of SRIF. SRIF is a potent regulator of endocrine function by inhibiting the release of growth hormone from the pitu-